Characterization of tumor cells and their differentiation from nonmalignant host cells present at the tumor site or infiltrating the tumor that may grow as colonies in an in vitro system is necessary to most efficiently analyze the effects of therapy using clonogenic cell assays. One goal of this proposal is the charaterization on nonmalignant cells and colonies from human brain tumors, in order to eliminate them from the evaluation of in vitro tumor cell survival studies and permit a more accurate representation of in situ tumor response. A panel of cell identificaton parameters consisting of histochemical stains, antigenic studies, functional criteria, and morphological characteristics will be used to identify normal host cells. Glial cells will be identified by the presence of glial fibrillary acidic protein and recognition by hybridoma secreted glioma-associated monoclonal antibodies. The cells disaggregated from tumor biopsies and cells growing in early passage monolayer culture, organ cultures, and xenografts will be analyzed. The neoplastic nature of cells will be suggested by tumor growth in immunodeprived mice, the lack of contact inhibition in monolayer cultures, abnormal chromosome counts, and, possibly, by monoclonal antibody techniques. The second goal of this proposal is the isolation and characterization of tumor cell clones from human brain tumors to gain increased knowledge of tumor heterogeneity and provide insights into the reasons for treatment failure. Improved methods for disaggregating single cells from solid tumor biopsies will be developed and studies will determine if the dissociated cells are representative of the cells within the tumor. Clonogenic cells will be considered tumor "stem" cells if self-renewal is demonstrated on serial subculture. Tumor cell clones will be isolated, characterized as outlined above and analyzed for cell kinetics and drug and radiation sensitivity. Tumor cell clones surviving antitumor treatment will be analyzed similarly.